Journal: Aging (Albany NY)

Article Title: Uhrf1 regulates H3K9me2 modification of mTOR to inhibit the effect of autophagy in myocardial ischemia-reperfusion injury

doi: 10.18632/aging.202722

Figure Lengend Snippet: Uhrf1 regulates mTOR expression through H3K9me2. ( A ) The relative mRNA expressions of Peak1-4 in Myocardial ischemia-reperfusion model in vitro were determined by qRT-PCR. ( B ) Western blot was used to detect the inhibitory effect of G9a inhibitor on H3K9me2 in cardiomyocytes by different concentrations (0.1 µM, 0.5 μM, 1.0 μM and 2.0 μM). GAPDH and the total histone H3 were used as loading controls. ( C ) The relative mRNA expressions of mTOR in myocardial ischemia-reperfusion model in vitro were determined by qRT-PCR after adding 1.0 μM G9a inhibitor to Si group. ( D ) Western blot was used to detect the expression level of mTOR protein after adding 1.0 μM G9a inhibitor to Si group. GAPDH serves as a loading control. ( E ) Expression of mTOR protein relative to GAPDH data from 3 biological repeats is shown. ( F ) The relative mRNA expressions of Peak1-4 in Myocardial ischemia-reperfusion model in vitro were determined by qRT-PCR after adding 1.0 μM G9a inhibitor to Si group. Data shown are mean ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. N=3 per group. Model, in vitro oxidative stress model; NC, negative control of RNAi; si, RNAi knockdown of Uhrf1; Uhrf1, Uhrf1 overexpression.

Article Snippet: After sealing with 5% skimmed milk, the PVDF membrane was incubated overnight with antibodies of Uhrf1, Bax, caspase-3, Bcl-2, Ki67 and mTOR from Bioss (Beijing, China), Beclin-1, P62, LC3 from Proteintech (Wuhan, China), p-mTOR and H3K9me2 from Immunoway (Newark, USA) and GAPDH from Goodhere Biotechnology (Hangzhou, China).

Techniques: Expressing, In Vitro, Quantitative RT-PCR, Western Blot, Control, Negative Control, Knockdown, Over Expression